1. Field of the Invention
The present invention provides methods for screening for substances which inhibit the oligomerization of NEMO and/or IKK-related complexes and/or signaling pathways based on the interference with NEMO oligomerization.
2. Description of the Background
NF-κB plays a prominent role in the inducible expression of genes involved in processes like immune and inflammatory responses, cell growth and death, oncogenesis (Hayden, M. S. and Ghosh, S. (2004) Genes Dev, 18, 2195-2224). In resting cells, NF-κB is sequestered in the cytoplasm through interaction with inhibitory proteins known as IκBs (Baldwin, A. S., Jr. (1996) Annu Rev Immunol, 14, 649-683). Stimuli transducing through the TNF, Toll-like, IL-1, B and T cell receptors as well as the HTLV-1 Tax protein promote IκB phosphorylation by IκB kinases (IKK), ubiquitination and degradation. This allows NF-κB to translocate into the nucleus where it activates gene transcription (Karin, M. and Ben-Neriah, Y. (2000) Annu Rev Immunol, 18, 621-663). IKK activation proceeds from outside the cell to the cytoplasm, but it can also occur from the nucleus to the cytoplasm (Wu et al (2006a) Nat Cell Biol). The NF-κB signaling cascade can be triggered through a <<classical>> or an <<alternative>> pathway (Bonizzi, G. and Karin, M. (2004) Trends Immunol, 25, 280-288). Unlike most proinflammatory stimuli which lead to transient increase in IKK activity, the oncoprotein Tax leads to a persistent activation of IKKs through both pathways (Sun, S. C. and Yamaoka, S. (2005) Oncogene, 24, 5952-5964).
In the classical pathway, IKK activity is associated with a multiprotein complex composed of two serine/threonine kinases, IKKα and IKKβ (Zandi et al (1997) Cell, 91, 243-252) and a non catalytic protein, NEMO (NF-κB essential modulator) (Yamaoka, S et al (1998) Cell, 93, 1231-1240). Cells lacking NEMO are unable to assemble the IKK complex and exhibit severe defects in NF-κB activation. The mechanism by which the IKK complex is activated remains unclear: trans-autophosphorylation (Tang et al (2003) J Biol Chem, 278, 37297-37305) as well as kinases such as MEKK3 (Yang et al (2001) Nat Immunol, 2, 620-624) or TAK1 (Takaesu et al (2003) J Mol Biol, 326, 105-115) have been implicated. Recently, the first genetic evidence that TAK1 was essential for TNFα and IL-1 NF-κB activation (Sato et al (2005) Nat Immunol, 6, 1087-1095) was published. Besides, converging evidence suggests that NEMO oligomerization also plays a crucial role in the IKK complex activation (Inohara et al., (2000) J Biol Chem 275, 27823-27831; Poyet et al (2000) J Biol Chem, 275, 37966-37977). The subunit composition and stoichiometry of the IKK complex is still debated although many reports suggest an association between the IKKα/IKKβ heterodimer and NEMO.